Evidence suggests that oestrogens are the major mitogens involved in promoting the growth of tumours in endocrine-dependent tissues, such as the breast and endometrium. Although plasma oestrogen concentrations are similar in women with or without breast cancer, breast tumour oestrone and oestradiol levels are significantly higher than in normal breast tissue or blood. In situ synthesis of oestrogen is thought to make an important contribution to the high levels of oestrogens in tumours and therefore inhibitors, in particular specific inhibitors, of oestrogen biosynthesis are of potential value for the treatment of endocrine-dependent tumours.
Over the past two decades, there has been considerable interest in the development of inhibitors of the aromatase pathway—which converts the androgen precursor androstenedione to oestrone. However, there is now evidence that the oestrone sulphatase (E1-STS) pathway, i.e. the hydrolysis of oestrone sulphate to oestrone (E1S to E1), as opposed to the aromatase pathway, is the major source of oestrogen in breast tumours. This theory is supported by a modest reduction of plasma oestrogen concentration in postmenopausal women with breast cancer treated by aromatase inhibitors, such as aminoglutethimide and 4-hydroxyandrostenedione, and also by the fact that plasma E1S concentration in these aromatase inhibitor-treated patients remains relatively high. The long half-life of E1S in blood (10-12 h) compared with the unconjugated oestrogens (20 min) and high levels of steroid sulphatase activity in liver and, normal and malignant breast tissues, also lend support to this theory.
PCT/GB92/01587 teaches novel steroid sulphatase inhibitors and pharmaceutical compositions containing them for use in the treatment of oestrone dependent tumours, especially breast cancer. These steroid sulphatase inhibitors are sulphamate esters, such as N,N-dimethyl oestrone-3-sulphamate and, preferably, oestrone-3-sulphamate (otherwise known as “EMATE”). EMATE has the following structure:

It is known that EMATE is a potent E1-STS inhibitor as it displays more than 99% inhibition of E1-STS activity in intact MCF-7 cells at 0.1 mM. EMATE also inhibits the E1-STS enzyme in a time- and concentration-dependent manner, indicating that it acts as an active site-directed inactivator. Although EMATE was originally designed for the inhibition of E1-STS, it also inhibits dehydroepiandrosterone sulphatase (DHA-STS), which is an enzyme that is believed to have a pivotal role in regulating the biosynthesis of the oestrogenic steroid androstenediol. Also, there is now evidence to suggest that androstenediol can be of even greater importance as a promoter of breast tumour growth. EMATE is also active in vivo as almost complete inhibition of rat liver E1-STS (99%) and DHA-STS (99%) activities resulted when it is administered either orally or subcutaneously. In addition, EMATE has been shown to have a memory enhancing effect in rats. Studies in mice have suggested an association between DHA-STS activity and the regulation of part of the immune response. It is thought that this can also occur in humans. The bridging O-atom of the sulphamate moiety in EMATE is important for inhibitory activity. Thus, when the 3-O-atom is replaced by other heteroatoms as in oestrone-3-N-sulphamate and oestrone-3-S-sulphamate, these analogues are weaker non-time-dependent inactivators.
Although optimal potency for inhibition of E1-STS can have been attained in EMATE, it is possible that oestrone can be released during sulphatase inhibition and that EMATE and its oestradiol congener can possess oestrogenic activity.
Ahmed et al (Biochem Biophys Res Commun Jan. 27, 1999;254(3):811-5) report on a structure-activity relationship study of steroidal and nonsteroidal inhibitors of STS.
The present invention seeks to provide novel compounds suitable for the inhibition of E1-STS as well as other therapeutic applications.
Additionally, estrogens undergo serial hydroxylation, methylation and conjugation reactions and recent evidence demonstrates that at least some of the products of these reactions possess unique biological activities independent of the classical oestrogen receptor (ER). 2-Methoxyestradiol (2-MeOE2) is a hydroxylation and methylation derivative of 17β-estradiol and has received considerable interest as an endogenous growth inhibitory and cytotoxic agent with potential use for cancer therapy.
2-MeOE2 inhibits the growth and/or promotes apoptosis of many malignant cell types, including ER positive and negative human breast cancer and leukaemic cell lines and primary chronic lymphocytic leukaemia cells. Oral administration of 2-MeOE2 inhibits the in vivo growth of transplanted human tumours in immunodeficient mice. In addition to its direct effects on the proliferation and survival of cancer cells, 2-MeOE2 also possesses significant anti-angiogenic activity.
We have generated a series of natural and synthetic oestrone and oestradiol derivatives to further examine the biological effects of this class of molecules and to identify compounds with enhanced activity. Of particular note, we showed that the 3-O-sulfamoylated derivatives of 2-methoxyestrone and 2-ethylestrone (2-methoxyestrone-3-O-sulfamate and 2-ethylestrone-3-O-sulfamate, respectively) were significantly more effective in inhibiting the growth and survival of breast cancer cells relative to their parental oestrones and 2-MeOE2 [Purohit et al 1999; MacCarthy-Morrogh et al 2000].
2-MeOE2 binds to the colchicine binding site on tubulin, and inhibits the polymerisation of tubulin dimers to give microtubules. 2-MeOE2 arrests cells in mitosis and induction of apoptosis is associated with phosphorylation of the anti-apoptotic BCL-2 protein. This is consistent with tubulin as a major target for 2-MeOE2 since other structurally diverse anti-tubulin agents, such as vincristine and paclitaxel have similar effects in cells. The enhanced activity of the 3-O-sulfamoylated oestrones was associated with more potent inhibition of tubulin polymerisation, enhanced mitotic arrest and apoptosis, and an increase in BCL-2 phosphorylation relative to parental compounds and 2-MeOE2.
More recently, 2-MeOE2 has been shown to inhibit the activity of both CuZn and Mn superoxide dismutases (SOD). SOD catalyse the breakdown of superoxide radical and are essential enzymes in protecting cells from reactive oxygen species (ROS). Cancer cells are thought to be particularly sensitive to inhibition of SOD, because of their enhanced production of superoxide ions and their reduced SOD expression. Inhibition of SOD activity and subsequent increase in ROS has been suggested to play a major role in induction of apoptosis by 2-MeOE2. In support of this idea Huang et al., showed that in addition to 2-MeOE2, 2-MeOE1 and 2-hydroxyestradiol, but not oestradiol or oestrone, also inhibited SOD and induced apoptosis. However, both of these compounds would also be expected to inhibit tubulin polymerisation to some extent and therefore this small series of compounds would not differentiate between these two modes of action. Therefore, it is not clear what are the key structural features of estrogens required for optimal activity.
The present invention seeks to provide novel compounds suitable for the inhibition of SOD as well as other therapeutic applications.